| IntrodutionIndividual identification regard individual as research object, including living body, corpse and biology material from of the human body, which examine the material, use theory and technology of forensic medicine and anthropolgic to detect,analyse,and confirm its individuals body source .The genetic markers detection is the most reliable and precise methods in individual identification, For the PCR technology is especially have high sensitivity , special character and simple operation, it is popular in detecting the DNA of the material. It has been developed from cell's level to the molecule level, grown from macroscopical level to the microcosmic level, has expanded the range in apph'cation of material evidence greatly, has improved the quality that forecsics material evidence appraises.But some cases often require us to examine the special material tissue what had been treated by paraffin wax and formalin fixation. For example,when the criminal is released on bail for medical treatment, he would offer the false pathology slices of organize, or others'malignant tumour pathology information; and the policyholder offers the fake pathological material while setting a claim, or in the pathology room, we often meet the mark mistake of the sample that is needed to confirm the tissues source of the material. To determine these special material in practice is more and more important .This research use high sensitive, special In situ PCR technology to determine the material which is treated by the paraffin wax and formalin fixation of different time, by detecting the genotype of MN blood group of the organizesslices. At the same time,we study the research of main influence factors , such as protease digestion,etc. We hope to set up a kind of steady, practical methods to detect the genotype of the material treated by paraffin wax and formlin, offer a kind of new detecting means for the forensic appraises and iditificition with individual material evidence.Material and methodsIn this study Sample blood 2ml, musculature 200mg each of 12 corpse, which had died within 48 hours, blood sample is using the methos of phenol / chloroform to draw DNA. Each of musculature tissue according to 10% formalin fixation time of 24h,48h,72h,3M dividing into groups separately,Embed by the paraffin wax, cut thick to into slices of 5 u.m, each sample make routine HE dye contrast.Use the agglutinate law directly to detect the red blood corpuscles MN phe-notype. Use routine PCR to examine the blood MN genotype of samples. The organization cuts into slices and examines by the in situ PCR, drip protease K 20 (xl with lOOmg/ml to digest respectively in pretreatment, increase with normal position positive cell account for total ratio of cell , according to the positive standard cells >75% , confirm the lightest digestion time, studying the influence and relationship of different fixation time with protease digesting each other , detecting the MN genotype of the organize slices at the same time .ResultWhether use the serum method, routine PCR and the in situ PCR means to examine 12 samples phenotype and genotype,the result is complete same, Of the 12 samples, 4 samples of the 1,6,7,11th are MM type,the 2,4,8,9,5,12th sample are NN type; the 3, 5, 10th sample are MN type .At the fixation time of 24h,48h,72h,and 3M,the most suitable protease digestion time of each group is separately 30.41 6.55min, 46. 67 6. 38min, 56. 25 8. 49min,58. 33 6. 38min. Statistics analyse indicate that the most suitableprotease digestion time of each group of different foxation time has remarkable difference (P <0.01).DiscussionIn practice of the forensic material evidence, one that is chemical disposal special material to examine, for example material treated by formalin fixation and paraffin wax is needed to distinguish the source to identificate. Because the formalin influence DNA to be drawn directly, fixation time is the important variation factor among them. In the course of drawing DNA by routine method , DNA i...
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